Our long-term goal is to understand how cell-cell interactions regulate the relative positions and proliferation of distinct muscle cell types during development, an issue of fundamental importance to the understanding of human birth defects, and diseases of muscle including muscular dystrophies and heart disease. We are using zebrafish to study the mechanisms controlling myogenic cell division and differentiation. A large number of zebrafish mutants with abnormal muscle patterning, or with muscle degeneration have recently been identified. These mutants, as well as the established methods for cell transplantations and molecular misexpression, provide powerful tools for understanding the mechanisms of myogenesis. We will test whether the proliferation of muscle precursors and their differentiation into specific fiber types is regulated by a series of inductive interactions from surrounding tissues. 1. We will characterize the role of notochord signaling in regulating the development of fiber type identity. We will define a genetic pathway for slow muscle development. 2. We will test the hypothesis that myogenic precursor cell division is regulated by signals from the notochord, including sonic hedgehog. We will examine the time course of myogenic precursor cell division and test whether myogenic precursor cell division is altered in mutants with impaired midline signaling. We will examine whether specific molecules in the Hh signaling pathway regulate cell proliferation. We will test whether cell proliferation and cell differentiation are linked by examining the effect of disrupting cell division on cell differentiation. 3. We will test the hypothesis that the timing of MRF expression regulates fiber type identity. We will test whether there is a strict correlation between the time of MRF expression and fiber type formed. We will use tissue specific, as well as inducible, promoters to express MRFs in paraxial cells that would normally generate fast muscle fibers, either before or after somite formation, and determine the fiber type these cells develop into. We will test whether forcing MRF expression in segmental plate cells is sufficient to rescue slow muscle development in mutants that are missing slow muscle fibers. 4. We will test the hypothesis that negative BMP4-like signals inhibit the development of specific fiber types. We will block cells from responding to BMP4-type proteins by expressing a dominant negative BMP4 receptor. We will ectopically activate BMP4 signaling by expressing an activated BMP4 receptor.